Cellular analysis of B cell activation in the rainbow trout.

Teleosts including the rainbow trout provide an interesting animal model, as they do not possess bone marrow or lymph node tissue. In trout, the anterior kidney is the main site for B lymphogenesis throughout life. Our lab uses expression patterns of B cell‐specific transcription factors as markers to investigate B cell development and activation in the trout. In mammalian species, transcription factors Pax5 and XbpI provide accurate markers for the terminal differentiation stages of B cells. Pax5 is expressed in mature and plasmablast stages but absent from plasma cells, while XbpI is not induced until the activated B cell stage. In this study we wished to investigate the activation stages of trout B cells using expression patterns for Pax5, XbpI, membrane and secreted immunoglobulin. In order to determine activation states in individual B cells, fixed and permeabilized trout immune cells were analyzed by flow cytometry using Pax5, XbpI, and Ig specific antibodies. This approach lead to the detection of three major B cell activation stages in trout immune tissues: the resting mature B cell stage, the activated B or plasmablast stage, and the plasma cell stage. Different immune tissues had different frequencies of each activation stage. While freshly isolated trout spleen and PBLs contained mostly resting mature B cells, anterior and posterior kidney tissues contained significant populations of Ig‐secreting cells. LPS‐activation lead to increased frequencies of activated B cells in spleen, but not PBL, while anterior and posterior kidney lose their Ig‐secreting cells. Our studies show for the first time that Pax5, XbpI, and IgM markers can be used in flow cytometric analyses to distinguish between resting B cells, activated B cells, and plasma cells of the rainbow trout. This research was funded through an AREA grant from the NIH.